1. Field of the Invention
This invention relates to improved methods and apparatus for electrophoretic analysis of samples, and more particularly to an improved method and apparatus providing completely automatic electrophoretic analyses.
2. Description of the Prior Art
Electrophoretic analysis is a well known technique for identifying substances, which is particularly suited for clinical analysis of constituents in blood serum. The technique is based on the fact that most substances contain particles which, when subjected to an electric field, will disperse in an identifiable pattern.
The technique is used to separate organic and inorganic compounds of human, animal and inanimate origin. In particular, separation of the components of various body fluids is of considerable diagnostic use. By this technique, it is possible to identify abnormal protein in fluids such as blood and urine. Additionally, various types of hemoglobin and isoenzymes can be separated electrophoretically.
Although electrophoretic techniques have considerable promise in the areas of medicine and industrial activity, present laboratory electrophoresis involves a number of manual steps which require a technician's time and which give rise to numerous errors and inaccuracies. These prior art electrophoretic techniques are illustratively described in U.S. Pat. No. 3,421,998. In that patent, electrophoretic separation involves the following steps:
1. An electrophoresis strip (porous strip) is saturated with a buffer solution and then placed in an electrophoresis chamber where it is fastened.
2. The serum sample to be tested is applied by a technician using a special sample applicator. The amount of sample applied is so small (about 1 microliter) that it is not quantitatively known so that the resulting determinations are of relative percentages of the protein fractions rather than absolute values.
3. Electrophoresis is then performed by applying a DC voltage for a period of time.
4. The electrophoresis strip is taken out of the chamber and treated in dye solutions, rinsing solutions and cleaning solutions by the technician.
5. The strip is then dried and the electrophoretic pattern is measured in an optical densitometer.
Thus, it is readily apparent that steps 1,2,4 and 5 involve a considerable number of manual operations. However, apparatus based on these operational steps is presently being extensively marketed.
Another type of electrophoresis operation is described in U.S. Pat. No. 3,432,414. In this reference, the analytical procedure consists of the following steps:
1. A liquid electrophoresis medium containing pH buffer and agarose is put into an electrophoretic vessel by a technician. This operation forms a thin layer on the top of a quartz plate which is transparent to UV radiation.
2. Each serum sample to be tested is brought into contact with a special sample applicator, as described in U.S. Pat. No. 3,360,454. The applicator is then brought into contact with the layer of electrophoretic medium. Here again, the amount of serum applied is not quantitatively known, hence the determination yields only relative percentages of fractions present in the serum.
3. After electrophoresis and quantization, the technician has to clear the electrophoretic chamber manually.
Again, this method of electrophoresis requires numerous manual steps which are time consuming and which lead to inaccuracies.
Other methods of electrophoresis are known which are not as widely used for clinical serum analysis. These other methods use columns such as are described in U.S. Pat. No. 3,384,564. However, this latter type of electrophoresis also involves several manual steps, as for instance:
1. The column is filled manually with gel, and the gel is then polymerized.
2. The sample to be tested is applied manually onto the gel in the column.
3. A buffer solution is added on top of the serum sample and electrophoresis is then carried out by the application of a voltage to the sample.
4. The gel column is then dyed and quantitatively measured. This procedure usually takes hours because the thick gel columns require a long time for dye diffusion to occur.
Thus, it is readily apparent that the prior art does not suggest an electrophoretic method or apparatus which can automatically yield determinations of absolute amounts of electrophoretic fractions. Further, the prior art does not teach or suggest an apparatus or method which is automatic or which can be automated in order to require only a minimum of operator time. In contrast, the present invention requires no manual handling of the electrophoretic medium or the sample to be analyzed, at any time during the analysis operation.
It will be appreciated that the manual operations described above are never performed at exactly the same time in the procedure. For example, the time between applying the sample and the start of the electrophoresis, or the time between the electrophoresis and the scanning of the developed pattern will vary slightly from one sample analysis to the next when manual steps are involved. This, in turn, affects the accuracy and reproducibility of results.
Accordingly, it is a primary object of this invention to provide completely automatic zone electrophoretic analysis.
It is another object of this invention to provide electrophoretic analysis which requires no manual steps and which utilizes improved apparatus in order to yield precision results quickly.
It is still another object of this invention to provide an electrophoretic technique for analysis of samples in which all steps of the technique are achieved with constant, reproducible timing for each sample which is analyzed.
It is still another object to provide an improved technique for electrophoretic analysis which reduces costs.
It is a further object of this invention to provide automatic electrophoresis with apparatus for improved scanning of electrophoretic patterns.
It is another object of this invention to provide an electrophoretic technique using an improved apparatus and method for loading sample serum for testing.
It is a still further object of this invention to provide improved electrophoretic techniques for determining isoenzymes.